Project Summary Diabetic retinopathy (DR) is a leading cause of vision loss, affecting about 93 million people worldwide. A major breakthrough in DR therapy is the recent approval of vascular endothelial growth factor (VEGF) inhibitors. However, anti-VEGF therapy has limited efficacy with a number of side effects. The critical barrier is how to identify other angiogenic targets for alternative or combination therapy of anti-VEGF-resistant DR. We have discovered a novel DR-associated angiogenic factor that selectively binds to and stimulates angiogenesis of diseased but not normal vessels. In contrast, VEGF binds to and induces the angiogenesis of both diseased and normal vasculature. This new ligand has a different receptor pathway than VEGF, and therefore their inhibitors will have different spectra of anti-angiogenic activity for alternative or combination therapy of DR. We developed a monoclonal antibody against this new target and demonstrated its capacity to alleviate DR in mice with high efficacy. Based on the normal phenotype of the knockout mice, we predicted and confirmed that the therapeutic antibody at an excessively high dose has minimal side effects. Therefore, anti-angiogenic therapy against this new ligand has the advantages of high efficacy, minimal side effects and a different mechanism of action than anti-VEGF. Antibody humanization is an essential step for the translation of this novel therapy. This project will humanize the monoclonal antibody (Aim 1). Humanized antibodies will be selected based on their ligand-binding affinity, neutralizing activity, therapeutic efficacy and side effects (Aim 2). Humanized antibody with optimal therapeutic activity and minimal side effects will be used for clinical trials in the next phase. Therefore, this project will lead to the development of a new drug for anti-angiogenic therapy of DR with high efficacy and safety.